The overall objective of the proposed research is to evaluate and refine a novel approach for optimizing the efficiency of human monoclonal antibody (MoAb) production, particularly MoAb with specificity for human tumor- associated antigens (TAA). Our general approach will be to generate human TAA-specific immune responses in situ with human lymphocytes that have been stably engrafted into C.B-17 scid mice (Mosier, et al., Nature 335:256, 1988). Three strategies are proposed: (a) to induce primary human antibody responses in scid mice engrafted with normal adult donor lymphocytes, (b) to stimulate secondary immune responses in scid mice engrafted with lymphocytes from tumor-bearing patients, and (c) to activate "hyperimmune" lymphocytes in scid mice engrafted with lymphocytes from cancer patients who have been actively immunized with tumor cells. The two types of tumors included in this proposal are breast cancer and malignant glioma. According, scid mice will be engrafted with lymphocytes from normal donors and from patients with breast cancer or malignant glioma, and the engrafted scid mice will be immunized with either purified breast carcinoma TAa glycoproteins (TAG12 or HER-2/neu) or an appropriate human tumor cell line (TAG12-positive T47-D and HER-2/neu-positive SK-BR-3 cells for breast carcinoma or the U-251MG glioma line used in a phase III active immunization trial for malignant glioma). Engrafted and multiply immunized scid mice will be monitored for serum levels of human immunoglobulins and human antibody reactivity with the respective immunogen. Immunized human lymphocytes will be retrieved from mice and used in fusions with B lymphoblastoid cell lines to generate hybridomas secreting human MoAb. Human MoAb will then be analyzed for binding specificities against human TAA and various normal and tumor tissues. Evaluation of this novel method of generating human MoAb derived from human lymphocytes hosted in scid mice may facilitate increased production of human MoAb for therapeutic applications in human malignancies and may yield new information on the antibody repertoire in human immune responses to tumors.